A novel leaf-movement inhibitor of a nyctinastic weed, Sesbania exaltata Cory, designed on a naturally occurring leaf-opening substance and its application to a potential, highly selective herbicide

Article abstract of DOI:10.1016/S0040-4039(02)01848-8

We isolated 4-O-β-D-glucopyranosyl-trans-p-coumarate (1), the leaf-opening substance of Sesbania exaltata Cory. The leaf-movement inhibitor, designed on the chemical mechanism of nyctinasty, could keep the leaves of S. exaltata open till the leaves withered and died.

Full text of DOI:10.1016/S0040-4039(02)01848-8

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 43 (2002) 7655–7658
A novel leaf-movement inhibitor of a nyctinastic weed,
Sesbania exaltata Cory, designed on a naturally occurring
leaf-opening substance and its application to a potential,
highly selective herbicide
a
a
b
a
a,
Noboru Takada, Eisuke Kato, Katsuhiro Ueda, Shosuke Yamamura and Minoru Ueda *
a
Department of Chemistry, Faculty of Science and Technology, Keio University, Hiyoshi, Yokohama 223-8522, Japan
b
Department of Chemistry, Biology and Marine Science, University of The Ryukyus, Nishihara-cho, Okinawa 903-0213, Japan
Received 7 August 2002; revised 29 August 2002; accepted 2 September 2002
Abstract—We isolated 4-O-b-D-glucopyranosyl-trans-p-coumarate (1), the leaf-opening substance of Sesbania exaltata Cory. The
leaf-movement inhibitor, designed on the chemical mechanism of nyctinasty, could keep the leaves of S. exaltata open till the
leaves withered and died. © 2002 Elsevier Science Ltd. All rights reserved.
We have isolated the biological substances controlling
nyctinasty from several plants, and revealed the chemi-
cal mechanism of control of nyctinasty by a biological
1
clock. Nyctinasty is controlled by two endogenous
leaf-movement factors of contrasting bioactivities: leaf-
closing substance which makes the leaf close and leaf-
opening substance which makes the leaf open. The
bioactivity of these leaf-movement factors is extremely
specific to the original plant from which they were
isolated. In the plant body, the rhythm of nyctinasty is
generated by change in balance of concentrations
between the two leaf-movement factors according to a
circadian rhythm. This change in the balance can be
attributed to hydrolysis of the glucoside-type leaf-
movement factor into the corresponding aglycon by the
substance. As an application, the leaf-movement
inhibitor can be used as a potential environment-
friendly herbicide of extremely high specificity to the
target weed. The leaf-movement inhibitor would have
complete selectivity to the target weed because of the
extremely high specificity of the leaf-opening substance
to the plant containing it. When we use the leaf-move-
ment inhibitor as a herbicide, it would be effective only
for the target leguminous weed and have no effect on
vicinal plants, insects, birds, animals, and human
beings. For example, genetically engineered soybean
2
action of b-glucosidase whose activity is controlled by a
3
biological clock. According to this mechanism, we
have succeeded in the inhibition of leaf-movement by
using a synthetic leaf-movement inhibitor designed on
the structure of a naturally occurring leaf-opening sub-
4
stance. And it was revealed that the nyctinastic plant
(
Monsanto’s Roundup Ready Soybeans) is widely culti-
vated in USA as the most important leguminous crop.
withers and dies without nyctinastic leaf-closure. This is
an important discovery to answer the historic question,
‘
Why does the leguminous plant sleep?’
Unfortunately, some leguminous weeds, such as Sesba-
nia exaltata Cory and Senna obtsusifolia, are resistant
to Roundup and can grow in the soybean fields after
the treatment of it. No existing herbicide can remove
these leguminous weeds without any damage to the
soybean, which belongs to the same genus. However,
leaf-movement inhibitor 5 for these weeds will be able
to remove them without any damage to the soybean
Also, this result showed that we can inhibit the leaf-clo-
sure of a voluntary leguminous plant using a synthetic
leaf-movement inhibitor designed on its leaf-opening
*
Corresponding author. Tel.: +81-45-566-1713; fax: +81-45-566-1697;
e-mail: ueda@chem.keio.ac.jp
0
040-4039/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(02)01848-8

7
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N. Takada et al. / Tetrahedron Letters 43 (2002) 7655–7658
and other vicinal organisms. In this paper, we describe
the isolation of a naturally occurring leaf-opening sub-
stance from Sesbania plant, and development of the leaf-
movement inhibitor of S. exaltata which is a potential
ecological herbicide.
HG-5 with 10% aqueous MeOH to give 4-O-b-D-
glucopyranosyl-trans-p-coumarate (1, 21 mg) as a
leaf-opening substance, whose structure was determined
6
by NMR and HRFABMS. Compound 1 showed leaf-
−
3
opening activity at 1.4×10 M against the leaves of
Sesbania plants, such as S. speciosa and S. exaltata. On
the other hand, 1 did not show any leaf-opening activity
against the leaves of nyctinastic plants belonging to
other genuses, such as Albizzia julibrissn Durazz.,
Leucaena leucocephala L., Phyllantus urinaria L. and
Desmanthus virgatus (Fig. 1). Therefore, it was clarified
that 1 was a genus-specific leaf-opening substance for
Sesbania plants.
Based on the previous study that the plants belonging to5
the same genus have the same leaf-movement factors,
we carried out the isolation of the leaf-opening substance
of Sesbania speciosa Taub belonging to the same genus
as one of the target weeds, S. exaltata.
The fresh whole plant of S. speciosa (14.9 kg), which was
collected in Okinawa, Japan, was extracted with MeOH
(
50 L) for a week and concentrated in vacuo. The con-
In the bioassay, natural leaf-opening substance 1 could
not make the leaves of S. exaltata wither and die,
because it may be easily metabolized by b-glucosidase in
the plant body before the leaves withered to die. The
artificial leaf-movement inhibitor, which is resistant to
centrated extract was partitioned with n-hexane, ethyl
acetate, and then n-BuOH. Isolation of the leaf-opening
substances was carried out based on a bioassay using a
leaf of S. speciosa. The bioactive fraction kept the leaf
open till 8:00 pm. The aqueous layer was separated by
Amberlite XAD-7 column chromatography eluted with
the hydrolysis by b-glucosidase, was necessary for the
development of the herbicide. In the previous study, it
was shown that a galactose derivative of glucoside-type
leaf-opening substance kept the leaves open until the
MeOH–H O (0:100, 10:90, 30:70, 50:50 and 100:0). The
2
leaf-opening activity was observed in the 10% aqueous
MeOH fraction. Thus, this fraction was separated by
MPLC using TSK-G3000S gel with 30% aqueous EtOH.
The bioactive fraction was further purified by HPLC
using preparative Develosil ODS-SR-5 column with 30%
aqueous MeOH, and then preparative Develosil ODS-
leaves withered and died because of its resistance to the
7
hydrolysis by b-glucosidase.
Therefore, we prepared several derivatives of 1 for the
structure–activity relationship study (Scheme 1).
Figure 1. Genus-specific leaf-opening activity of 1. (a) S. speciosa Taub., (b) S. exaltata Cory, (c) A. julibrissn Durazz., (d) L.
−
3
leucocephala L.,(e) P. urinaria L., (f) D. virgatus. Every leaf was treated with 3×10 M of 1. This photo was taken at 8:00 pm.
Scheme 1. Synthesis of 4-O-D-galactopyranosyl-trans-p-coumarate (5).

N. Takada et al. / Tetrahedron Letters 43 (2002) 7655–7658
7657
Figure 2. The structure–activity relationship study on the leaf-opening substance of S. exaltata.
Namely, a coupling reaction of methyl trans-p-cou-
marate (2) and -acetobromogalactose 3 was carried
out with AgOTf–MS 4A to give the desired -galac-
toside 4a (52%) and undesired a-galactoside 4b (29%).
Desired 4a was deprotected using aqueous KOH to give
inhibitor 5 would be a potential, highly selective herbi-
cide of S. exaltata.
D
b
In conclusion, we have succeeded in the isolation and
structural determination of the leaf-opening substance,
4-O-b-D-glucopyranosyl-trans-p-coumarate (1), which
is effective against S. exaltata Cory, a leguminous weed
growing in a field of genetically engineered soybean
after the treatment of Roundup. Based on our pro-
posed mechanism of the leaf-movement in leguminous
plants, we designed and prepared the leaf-movement
inhibitor 5. This synthetic inhibitor showed genus-spe-
cific activity against genus Sesbania, and was not
5
(90%). Other derivatives 6 and 7 were also prepared
8
by similar procedure. The sugar derivatives 5 and 6
were as effective as 1 against the leaves of S. exaltata,
while 7, which was the cis isomer of 1, was not effective
−
2
even at 1.4×10 M in the bioassay. Based on the
structure–activity relationship study using some deriva-
tives of 1, it was suggested that the structure of aglycon
was essential for leaf-opening activity, while the struc-
ture of the sugar moiety was not important (Fig. 2).
However, it was suggested that the presence of the
sugar part was necessary for leaf-opening activity, since
trans-p-coumarate did not show any leaf-opening activ-
ity. These results suggested that 5 and 6, which would
metabolized by
b-glucosidase to make the Sesbania
leaves wither and die. This result shows that the leaf-
movement inhibition designed on the leaf-opening sub-
stance can be used as a potential, highly selective
herbicide without damage to the surrounding
organisms.
not be hydrolyzed by b-glucosidase, can be used as a
potential herbicide for S. exaltata.
Then, we carried out the bioassay using 5 against the
leaves of S. exaltata. Fortunately, 5 kept the leaves
open till the leaves withered and died on the bioassay
Acknowledgements
This work was supported by the Ministry of Education,
Science, Sports and Culture (Japan) for Grant-in-Aid
for Scientific Research (No. 1302467 and 12680598),
Pioneering Research Project in Biotechnology given by
the Ministry of Agriculture, Forestry and Fisheries,
Kato Memorial Bioscience Foundation, Naito Founda-
tion, Kurata Foundation and Moritani Foundation.
(
Fig. 3). This result suggested that the leaf-movement
References
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2
3
4
. Ueda, M.; Yamamura, S. Angew. Chem., Int. Ed. 2000, 39,
1400–1414 and references cited therein.
. Ueda, M.; Asano, M.; Sawai, Y.; Yamamura, S. Tetra-
hedron 1999, 55, 5781–5792.
. Ohnuki, T.; Ueda, M.; Yamamura, S. Tetrahedron 1998,
Figure 3. The status of leaves of S. exaltata at 8:00 pm by
treatment with 4×10 M of leaf-movement inhibitor 5. (a) 1st
day; (b) 2nd day; (c) 3rd day; (d) control.
54, 12173–12184.
−
3
. Ueda, M.; Sugimoto, T.; Sawai, Y.; Yamamura, S. Tetra-
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7
658
N. Takada et al. / Tetrahedron Letters 43 (2002) 7655–7658
1
5
6
. Ueda, M.; Okazaki, M.; Ueda, K.; Yamamura, S. Tetra-
4-O-a-
D
-Mannnopyranoyl-trans-p-coumarate (6):
H
hedron 2000, 56, 8101–8105.
NMR (400 MHz, D O): 7.34 (2H, d, J=8.6 Hz), 7.13 (1H,
d, J=16.0 Hz), 6.93 (2H, d, J=8.6 Hz), 6.20 (1H, d,
J=16.0 Hz), 4.93 (1H, d, J=2.0 Hz), 3.96 (1H, dd, J=2.0,
2
1
. 4-O-
b
-
D
-Glucopyranosyl-trans-p-coumarate (1): H NMR
(
400 MHz, D O): 7.41 (2H, d, J=8.8 Hz), 7.17 (1H, d,
2
J=15.6 Hz), 6.95 (2H, d, J=8.8 Hz), 6.25 (1H, d, J=15.6
Hz), 4.98 (1H d, J=7.6 Hz), 3.75 (1H, dd, J=2.0, 12.4
Hz), 3.57 (1H, dd, J=5.6, 12.4 Hz), 3.47 (1H, ddd, J=2.0,
3
.4 Hz), 3.85 (1H, dd, J=3.4, 9.4 Hz), 3.56 (1H, dd,
J=3.7, 6.5 Hz), 3.54 (1H, dd, J=2.0, 6.5 Hz), 3.49 (1H,
dd, J=3.7, 5.4 Hz), 3.47 (1H, ddd, J=2.0, 5.4, 9.4 Hz)
ppm; C NMR (100 MHz, D O): 176.7, 157.2, 141.1,
1
6
5
.6, 9.0 Hz), 3.43 (1H, dd, J=9.0, 9.2 Hz), 3.39 (1H dd,
13
2
13
J=7.6, 9.0 Hz), 3.31 (1H, dd, J=9.0, 9.2 Hz) ppm;
C
30.6, 130.1 (2C), 123.6, 117.9 (2C), 98.8, 74.3, 71.3, 70.8,
NMR (100 MHz, D O): 176.6, 158.3, 141.1, 131.0 130.2
2
7.5, 61.6 ppm; IR (film) w 3267, 1638, 1604, 1551, 1509
(
2C), 123.7, 117.5 (2C), 100.7, 77.0, 76.4, 73.7, 70.3, 61.3
−1
20
D
cm ; [h] +98.6° (c 1.00, H O); HR FAB MS (negative):
2
−
1
21
ppm; IR (film) w 3359, 1640, 1604, 1509 cm ; [h] −59.2°
(
−
D
[M−H] found m/z 325.0929, C H O requires m/z
−
15 17 8
c 1.00, H O); HR FAB MS (negative): [M−H] found m/z
2
3
4
25.0923.
-O- -Glucopyranoyl-cis-p-coumarate (7):
3
25.0938, C H O requires m/z 325.0923.
15 17 8
1
b
-
D
H NMR
7
. Ueda, M.; Sawai, Y.; Yamamura, S. Tetrahedron 1999, 55,
0925–10936.
. 4-O- -Galactopyranoyl-trans-p-coumarate
(
400 MHz, D O): 7.23 (2H, d, J=9.0 Hz), 6.89 (2H, d,
2
1
J=9.0 Hz), 6.29 (1H, d, J=12.8 Hz), 5.81 (d, J=12.8 Hz),
1
8
b
-
D
(5):
H
4
3
9
.96 (1H, d, J=7.4 Hz), 3.73 (1H, dd, J=2.4, 12.6 Hz),
.56 (1H, dd, J=5.5, 12.8 Hz), 3.45 (1H, ddd, J=2.4, 5.5,
.4 Hz), 3.42 (1H, dd, J=9.1, 9.3 Hz), 3.37 (1H, dd,
NMR (270 MHz, D O): 7.40 (2H, d, J=8.5 Hz), 7.17 (1H,
2
d, J=16.1 Hz), 6.95 (2H, d, J=8.5 Hz), 6.23 (1H, d,
J=16.1 Hz), 4.89 (1H d, J=7.0 Hz), 3.81 (1H, d, J=2.8
13
13
J=7.4, 9.3 Hz), 3.30 (1H, dd, J=9.1, 9.4 Hz) ppm;
NMR (100 MHz, D O): 178.5, 157.8, 132.6, 132.4 (2C),
1
C
Hz), 3.72–2.55 (5H, m) ppm; C NMR (100 MHz, D O):
2
1
1
1
77.9, 158.5, 141.1, 130.9, 130.2 (2C), 123.6, 117.5 (2C),
2
31.4, 126.8, 117.8 (2C), 101.6, 77.7, 77.1, 74.5, 71.0, 62.1
01.3, 76.3, 73.4, 71.3, 69.3, 61.6 ppm; IR (film) w 3316,
−
1
22
D
−
1
20
D
ppm; IR (film) w 3400, 1700, 1560, 1400 cm ; [h] −40.2°
637, 1604, 1508 cm ; [h] −46.7° (c 1.00, H O); HR
2
−
−
(
c 0.25, H O); HR FAB MS (negative): [M−H] found m/z
2
FAB MS (negative): [M−H] found m/z 325.0938,
C H O requires m/z 325.0923.
325.0936, C15H O requires m/z 325.0923.
17 8
15
17
8